BRAF Molecular Testing Research Articles

BRAFV600E immunohistochemistry can reliably substitute BRAF molecular testing in the Lynch syndrome screening algorithm in colorectal cancer.

The Lynch syndrome (LS) screening algorithm requires BRAF testing as a fundamental step to distinguish sporadic from LS-associated colorectal carcinomas (CRC). BRAF testing by immunohistochemistry (IHC) has shown variable results in the literature. Our aim was to analyse concordance between BRAFV600E IHC and BRAF molecular analysis in a large, mono-institutional CRC whole-slide, case series with laboratory validation. MisMatch repair (MMR) protein (hMLH1, hPMS2, hMSH2, and hMSH6) and BRAFV600E IHC were performed on all unselected cases of surgically resected CRCs (2018-2023). An in-house validation study for BRAFV600E IHC was performed in order to obtain optimal IHC stains. BRAFVV600E IHC was considered negative (score 0), positive (scores 2-3), and equivocal (score 1). Interobserver differences in BRAFV600E IHC scoring were noted in the first 150 cases prospectively collected. Nine-hundred and ninety CRCs cases (830 proficient (p)MMR/160 deficient (d)MMR) were included and all cases performed BRAFV600E IHC (BRAFV600E IHC-positive 13.5% of all series; 66.3% dMMR cases; 3.4% pMMR cases), while 333 also went to BRAF mutation analysis. Optimal agreement in IHC scoring between pathologists (P < 0.0001) was seen; concordance between BRAFV600E IHC and BRAF molecular analysis was extremely high (sensitivity 99.1%, specificity 99.5%; PPV 99.1%, and NPV 99.5%). Discordant cases were reevaluated; 1 score 3 + IHC/wildtype case was an interpretation error and one score 0 IHC/mutated case was related to heterogenous BRAFV600E IHC expression. Among the 12 IHC-equivocal score 1+ cases (which require BRAF molecular analysis), three were BRAF-mutated and nine BRAF-wildtype. BRAFV600E IHC can be used as a reliable surrogate of molecular testing after stringent in-house validation.

BRAF V600E Mutations and Beyond: A Molecular Perspective of Melanoma from a Tertiary Cancer Referral Center of India

Objectives Malignant melanoma demonstrates frequently occurring mutations of genes in the serine/threonine kinase pathway, namely BRAF, NRAS, and neurofibromin 1. There is rare documentation of a detailed analysis of these mutations in cases of melanoma among Indian patients. We present molecular features in cases of malignant melanoma, diagnosed at a tertiary cancer referral center in India, over a period of 8 years (2011–2018). Materials and Methods This study was performed on formalin fixed paraffin embedded tissues of 88 histologically confirmed cases of malignant melanoma. BRAF gene alterations were studied by both Sanger sequencing and real-time polymerase chain reaction techniques (n = 74). Molecular testing for BRAF and NRAS gene alterations was accomplished in 74/88 cases (80%). Molecular test results were correlated with clinicopathological features using IBM SPSS Statistical software 25.0. Results The age ranged from 13 to 79 years (median = 57), with a M:F ratio of 1.4:1. BRAF mutations were observed in 12/74 (16.21%) patients, including V600E (n = 7), A594T (n = 1), T599 = (n = 2), V600K (n = 1), and Q612P (n = 1), while NRAS mutations were observed in 6/38 (15.7%) patients. Among various subtypes, nodular melanoma was the most frequent subtype (33%) among cutaneous malignant melanomas. Among non-cutaneous melanomas, mucosal melanomas were observed in 37.5% of cases. Conclusion This constitutes one of the few reports on comprehensive analysis of molecular alterations underlying melanomas in Indian patients. A larger sample size, with more extensive molecular markers, would yield additional information on the disease manifestation.

Turnaround Times in Melanoma BRAF Testing and the Impact on the Initiation of Systemic Therapy at a Single Tertiary Care Cancer Center.

The identification of BRAF mutations in melanoma enables targeted therapy and improves patient outcomes. Barriers to BRAF molecular testing affect the quality of care and therapeutic options. This retrospective study mapped BRAF testing timelines in adult patients with melanoma at the Princess Margaret Cancer Centre to identify obstacles to timely BRAF reporting and its impact on the initiation of therapy. Sixty-six cases were included. The median time between BRAF request and result was 12 days (95% CI, 8 to 15) when the BRAF test was ordered by pathology, compared with 20 days (95% CI, 16 to 23) if the test was requested by another specialist (P < .001). When the BRAF test and biopsy were performed within the same institution, the BRAF median turnaround time (TAT) was 13 days (95% CI, 6 to 19) compared with 19 days (95% CI, 16 to 21) if the sample was transferred from another institution (P = .02). Forty-seven patients received systemic therapy, and 20 had metastatic disease. In the metastatic subgroup, if the BRAF result was available at the first medical oncology visit, the initiation of treatment was 20 days (95% CI, 9.6 to 30.3), but was delayed to 31 days (95% CI, 10.8 to 51.1) if the BRAF result was not available (P = .03). This study showed variations in BRAF test results in TAT. One factor affecting this timeline is the transfer time, which can be streamlined by pathology reflex testing. Delays in TAT affect the timing and type of therapeutic intervention, especially in patients with stage IV disease.

A rare case of erdheim-chester disease with braf mutation – a case report

Erdheim-Chester disease (ECD) is a rare histiocytic neoplasm with multisystemic involvement, commonly involving the bones, cardiovascular organs and retroperitoneal region. We report a 68-year-old female who was admitted due to a fall following an episode of dizziness, and was noted to have a retroperitoneal soft tissue mass involving the bilateral perinephric spaces and bowel mesentery, which extended superiorly into the thoracic cavity, causing encasement of the ureters, adrenal glands and major vessels. A biopsy revealed a histiocytic infiltrate with foamy cytoplasm, which was positive for CD163, fascin and factor XIIIa on immunohistochemical staining, but was negative for CD1a, S100, AE1/3 and HMB45. Molecular testing for BRAF was positive. The overall clinical, imaging and histology findings were supportive of a diagnosis of ECD. Awareness of this entity and exclusion of its differential diagnoses is important to ensure correct diagnosis for subsequent targeted therapy. 1.Brousse N, Pileri SA, Haroche J, et al. Erdheim Chester disease. In: Swerdlow SH, Campo E, Harris NL, et al. WHO Classification of Tumours of Haematopoietic and Lymphoid Tissue. Revised 4th ed. Lyon: IARC, 2017; 481–2.2.Goyal G, Heaney ML, Collin M, et al. Erdheim-Chester disease: consensus recommendations for evaluation, diagnosis, and treatment in the molecular era. Blood 2020; 135: 1929–45.3.Aubart FC, Emile JF, Carrat F, et al. Targeted therapies in 54 patients with Erdheim-Chester disease, including follow-up after interruption (the LOVE study). Blood 2017; 130: 1377–80.

BRAF testing timelines and impact on the starting of systemic treatment.

e21575 Background: Targeted therapy with BRAF and MEK inhibitors constitute part of the standard treatment for BRAF V600 mutated melanoma. Timely detection of BRAF mutation is necessary for clinicians and patients to make treatment decisions. We aimed to map the BRAF testing timelines from the time of request until the reported result in order to assess obstacles to timely BRAF reporting in our community, and its impact on the initiation of systemic therapy. Methods: In this single-center retrospective study, we included adult patients referred to the Medical Oncology Department at the Princess Margaret Cancer Centre (PM) from January 2019 to August 2019 with histologically confirmed cutaneous or mucosal melanoma and BRAF molecular testing performed in 2019. The Log-Rank method was applied to detect differences in BRAF turnaround time and time to treatment initiation in specified subgroups. A p value &lt; 0.05 was considered statistically significant. Results: Sixty-six cases were identified. The median age was 64 (24-88), and 42 (64%) were male. At the time of BRAF request, 10 (15%) patients had stage II, 33 (50%) had stage III, and 23 (35%) had stage IV disease. Twenty-eight (43%) were positive for the BRAF V600E/K mutation by ARMS assay and 4 (6%) for other variants by NGS test. Thirty-three (50%) patients had the BRAF test available at their first PM Medical Oncology visit. Median time between BRAF request and result was 17 days; when a reflex BRAF test was ordered by the Pathology Department, the median turnaround time was 12 days (95% CI 8-15), compared to 20 days (95% CI 16-23) if the order was requested by another specialist ( p &lt; 0.001). Median time to transfer samples between institutions was 6 days. If the BRAF test was processed within the institution where the biopsy was performed, the BRAF median turnaround time was 13 days (95% CI 6-19) compared to 19 days (95% CI 16-21) if a sample was transferred to another institution ( p = 0.02). In total, 49 patients had systemic therapy. Median time between the first visit with Medical Oncology and treatment initiation was 29 days and was not statistically different if the BRAF result was available or not (28 vs. 34 days; p = 0.09). In the subgroup with stage IV disease (Table), 20 patients received systemic therapy; the median time to treatment initiation was 24 days and differed with BRAF result availability (20 vs. 31 days, p = 0.03). Conclusions: The current BRAF testing timeline at the PM varies from days to weeks. A major factor impacting this timeline is transfer time, which can be streamlined by pathology reflex testing. Delays in turnaround time appears to impact subsequent timing and type of therapeutic interventions, especially in patients with stage IV disease.[Table: see text]

BRAF molecular testing in cytopathology: Implications for diagnosis, prognosis, and targeted therapeutics.

BRAF molecular testing in cytopathology: Implications for diagnosis, prognosis, and targeted therapeutics.

Hybrid Capture-Based Genomic Profiling Identifies BRAF V600 and Non-V600 Alterations in Melanoma Samples Negative by Prior Testing

BRAF and MEK inhibitors are approved for BRAF V600-mutated advanced melanoma, with response rates of up to 70%. Responses to targeted therapies have also been observed for diverse non-V600 BRAF alterations. Thus, sensitive, accurate, and broad detection of BRAF alterations is critical to match patients with available targeted therapies. Pathology reports were reviewed for 385 consecutive melanoma cases with BRAF mutations or rearrangements identified using a hybrid capture-based next-generation sequencing comprehensive genomic profiling (CGP) assay during the course of clinical care. Records of prior BRAF molecular testing were available for 79 (21%) cases. Of cases with BRAF V600 mutations, 11/57 (19%) with available data were negative by prior BRAF testing. Prior negative BRAF results were also identified in 16/20 (80%) cases with non-V600 mutations, 2 of which harbored multiple BRAF alterations, and in 2/2 (100%) cases with activating BRAF fusions. Clinical outcomes for a subset of patients are presented. CGP identifies diverse activating BRAF alterations in a significant fraction of cases with prior negative testing. Given the proven clinical benefit of BRAF/MEK inhibitors in BRAF-mutated melanoma, CGP should be considered for patients with metastatic melanoma, particularly if other testing is negative. Published guidelines for melanoma treatment recommend BRAF mutational analysis, but little guidance is provided as to selection criteria for testing methodologies, or as to clinical implications for non-V600 alterations. This study found that hybrid capture-based next-generation sequencing can detect BRAF alterations in samples from a significant fraction of patients with advanced melanoma with prior negative BRAF results. This study highlights the need for oncologists and pathologists to be critically aware of coverage and sensitivity limitations of various assays, particularly regarding non-V600E alterations, of which many are potentially targetable.

Prognostic significance of BRAF mutation alone and in combination with microsatellite instability in stage III colon cancer.

The prognostic significance of biomarkers in colorectal cancer is still being defined. This study aimed to determine the prognostic significance of BRAF mutation alone and in combination with microsatellite instability (MSI), in stage III colon cancer. Curatively resected stage III colon cancers were studied from a 33-year period. Clinicopathological data were collated (adjuvant chemotherapy, age, gender, obstruction, perforation, tumour location, grade, presence of mucin, nodal stage, extramural vascular, and perineural invasion). MSI status was established and molecular testing for BRAF (V600E) was performed. Four mutation categories were examined: "traditional" (microsatellite stable [MSS]/BRAF -ve), "presumed Lynch" (MSI/BRAF -ve), "sporadic MSI" (MSI/BRAF +ve), and "other BRAF" (MSS/BRAF +ve). These factors were correlated with cancer-specific survival. In total, 686 unselected cases met our inclusion criteria, of which 15.7% had a BRAF mutation and 13.8% showed MSI. In the adjusted analysis, neither BRAF mutation nor MSI mutation were independently prognostic. On univariate analysis, survival in presumed Lynch cancers was similar to traditional cancers (5-year survival: 62% and 61%, respectively). While there was no difference in cancer-specific survival between sporadic MSI and other BRAF, both these tumour group had poorer outcome when compared to traditional or presumed Lynch cancers. Adjusted analysis of the four groups, however, showed that none of the subgroups were independently prognostic. BRAF-mutated cancers demonstrated a trend toward poorer outcomes, however, when adjusted for clinicopathological factors and chemotherapy, BRAF mutation was not found to be an independent prognostic biomarker in stage III colon cancer, even when combined with MSI.

1234P - Hybrid-capture based genomic profiling identifies BRAF V600 and non-V600 alterations in melanoma samples negative by prior testing

Background: BRAF and MEK inhibitors are approved for V600-mutated melanoma, and response rates of up to 70% are seen for patients with V600 mutations. Responses to targeted therapies have also been observed for a variety of non-V600 BRAF alterations. Thus, sensitive, accurate, and broad detection of BRAF alterations is critical to match patients with available targeted therapies. Methods: Pathology reports were reviewed for 385 consecutive melanoma cases (Mar 2016 - Mar 2017) with BRAF mutations or rearrangements identified using a hybrid-capture based next generation sequencing (NGS) assay during the course of clinical care. Results: Records of prior BRAF molecular testing were available for 79 (21%) cases, utilizing PCR (n = 30), Sanger sequencing (n = 13), IHC (n = 10), non-hybrid capture based NGS (n = 9), or other or unspecified methodology (n = 17). Of cases with BRAF V600 mutations 11/57 (19%) with available data were negative by prior BRAF testing, including 2/11 (18%) with confirmation that the same biopsy was tested. In cases with BRAF V600 mutations, there was no significant difference in mutant allele frequencies (median 35% vs. 40%, p = 0.25) or percentage of tumor nuclei (median 50% for both, p = 0.97) between samples with prior negative and prior positive results. Prior negative results were also identified in 16/20 (80%) cases with non-V600 mutations, two of which harbored multiple BRAF alterations [K601E (4), D594A/G/N (4), S467L (2), L584F (2), G464V, G466V, G469V, E586K, N581I, L597Q, A589_T599insT]. Two of 2 (100%) cases with activating BRAF fusions also had prior negative BRAF results. Clinical outcomes for a subset of patients will be presented. Conclusions: Despite approved companion diagnostics, significant variability exists in methods for BRAF testing in the clinical setting. Hybrid-capture based NGS identifies diverse activating mutations and fusions, including BRAF V600E, in a significant fraction of cases for which prior BRAF testing returned negative results. Given the proven clinical benefit in patients with BRAF alterations treated with match targeted therapies, hybrid-capture based NGS should be considered for patients with metastatic melanoma, particularly if other testing is negative. Legal entity responsible for the study: Foundation Medicine, Inc. Funding: Foundation Medicine, Inc. Disclosure: A. Wang, J.S. Ross, P.J. Stephens, S.M. Ali, A.B. Schrock, V.A. Miller: Employee with stock ownership in Foundation Medicine, Inc. All other authors have declared no conflicts of interest.

Detection of BRAF mutations on direct smears of thyroid fine-needle aspirates through cell transfer technique.

To determine the utility of the cell transfer technique (CTT) for BRAF molecular testing on thyroid fine-needle aspiration (FNA) specimens. Polymerase chain reaction (PCR)-based BRAF molecular testing was performed on tissues obtained through CTT from both air-dried and ethanol-fixed direct smears of thyroid FNA specimens and then compared with the corresponding thyroidectomy formalin-fixed, paraffin-embedded (FFPE) tissues on 30 cases. BRAF testing was successfully performed on 29 of 30 air-dried CTT, 27 of 30 ethanol-fixed CTT, and 27 of 30 FFPE tissues. The results exhibited 11, 13, and 13 BRAF mutations and 18, 14, and 14 wild types for the air-dried CTT, the ethanol-fixed CTT, and the FFPE tissues, respectively. The concordance rate was 96% between air-dried and ethanol-fixed CTT tissues, 88% between air-dried CTT and FFPE tissues, and 92% between ethanol-fixed CTT and FFPE tissues. PCR-based BRAF mutational testing can be reliably performed on the direct smears of the thyroid FNA specimens through the application of CTT.

Detection of BRAF Mutation in Metastatic Melanoma Utilizing Cell-Transferred Cytological Smears

Objectives: Fine-needle aspiration (FNA) is frequently used to diagnose metastatic melanoma. In this study, we validated the use of cell-transferred cytological smears for BRAF molecular testing. Study Design: We conducted a search of our laboratory information system for the period 2011-2013 in order to identify surgical pathology cases of either primary or metastatic melanomas in which BRAF mutation analyses had already been performed. Thirty FNA cases with diagnoses of metastatic melanoma from the same patients were identified. Direct smears from each FNA case were selected for mutation analyses using the cell transfer technique. Results: Mutation analyses were successfully performed on 28 of 30 FNA cases (93%) using the cell-transferred cytological smears. In 25 cases (8 BRAF mutations and 17 BRAF wild types), there was 100% agreement for the BRAF mutation between the cell-transferred cytological smears and the formalin-fixed paraffin-embedded tissues. Three FNA cases showed BRAF mutations that had not been detected in the correlated surgical specimens which were tested twice, and 2 cases failed to work. Conclusions: Cell-transferred cytological smears are a reliable and alternative resource for detecting BRAF mutations in metastatic melanoma.

Detection of BRAF Mutations on Direct Smears of Thyroid Fine Needle Aspirates Using the Cell Transfer Technique

Introduction: A mutation of the BRAF gene is the most common genetic alteration in papillary thyroid carcinoma (PTC). Molecular techniques that can identify BRAF mutations could be an important ancillary tool in the diagnosis of thyroid carcinoma, especially in cases of indeterminate thyroid fine needle aspirates (FNA). The cell transfer technique (CTT) has proven to be useful for obtaining cells for molecular testing from FNA direct smears. Materials and Methods: PCR-based BRAF molecular testing was performed on one air-dried and one ethanol-fixed CTT smear prepared from a thyroid FNA and on sections obtained from the corresponding thyroidectomy formalin-fixation paraffin embedded (FFPE) blocks from 31 cases diagnosed by histopathologic assessment as classical PTC (18), follicular variant PTC (4), follicular carcinoma (1), follicular adenoma (1) and benign non-neoplastic lesions (7). Results: BRAF testing was successfully performed on 30/31 air-dried CTT (97%), 28/31 ethanol-fixed CTT (90%) and 28/31 FFPE (90%). The results included 12 BRAF mutations and 18 wild types (WT) for the air-dried CTT, 14 BRAF mutations and 14 WT for the ethanol-fixed CTT and 13 BRAF mutations and 15 WT for the FFPE. The concordance rate between air-dried and ethanol-fixed CTT was 96% and was 86% between CTT and FFPE. There were 2 mutations detected on CTT which were WT on the corresponding FFPE. Conversely, 2 mutations identified on the FFPE were WT on the corresponding CTT. Conclusions: Using the FFPE as the gold standard, the sensitivity and specificity for detection of BRAF mutation on the CTT direct smears were 85% and 87%, respectively. BRAF mutation is a highly specific marker for thyroid carcinoma. For those patients with indeterminate thyroid FNA results, the use of CTT to select atypical cells for the BRAF mutation analysis should enhance the diagnostic sensitivity for thyroid FNA and reduce the need for repeat aspirates.

A clinical algorithm for fine-needle aspiration molecular testing effectively guides the appropriate extent of initial thyroidectomy.

To test whether a clinical algorithm using routine cytological molecular testing (MT) promotes initial total thyroidectomy (TT) for clinically significant thyroid cancer (sTC) and/or correctly limits surgery to lobectomy when appropriate. Either TT or lobectomy is often needed to diagnose differentiated thyroid cancer. Determining the correct extent of initial thyroidectomy is challenging. After implementing an algorithm for prospective MT of in-house fine-needle aspiration biopsy specimens, we conducted a single-institution cohort study of all patients (N = 671) with nonmalignant cytology who had thyroidectomy between October 2010 and March 2012, cytological diagnosis using 2008 Bethesda criteria, and 1 or more indications for thyroidectomy by 2009 American Thyroid Association guidelines. sTC was defined by histological differentiated thyroid cancer of 1 cm or more and/or lymph node metastasis. Cohort 2 patients did not have MT or had unevaluable results. In cohort 1, MT for a multigene mutation panel was performed for nonbenign cytology, and positive MT results indicated initial TT. MT guidance was associated with a higher incidence of sTC after TT (P = 0.006) and a lower rate of sTC after lobectomy (P = 0.03). Without MT results, patients with indeterminate (follicular lesion of undetermined significance/follicular or oncocytic neoplasm) cytology who received initial lobectomy were 2.5 times more likely to require 2-stage surgery for histological sTC (P < 0.001). In the 501 patients with non-sTC for whom lobectomy was the appropriate extent of surgery, lobectomy was correctly performed more often with routine preoperative MT (P = 0.001). Fine-needle aspiration biopsy MT for BRAF, RAS, PAX8-PPARγ, and RET-PTC expedites optimal initial surgery for differentiated thyroid cancer, facilitating succinct definitive management for patients with thyroid nodules.

Macrofollicular Variant of Papillary Thyroid Carcinoma with Extensive Lymph Node Metastases

The macrofollicular variant of papillary thyroid carcinoma is a rare subtype of the follicular variant of papillary thyroid carcinoma and is usually characterized by an indolent clinical course. The tumors are prone to be misdiagnosed as benign due to their macrofollicular architecture and bland cytologic features. We report a rare case of the macrofollicular variant of papillary thyroid carcinoma with extensive lymph node metastases. The patient was a 48-year-old female with a right thyroid nodule and multiple enlarged lymph nodes in the right neck. It was not possible to make a definitive diagnosis of malignancy on fine-needle aspiration cytology and intraoperative frozen section. She underwent total thyroidectomy with right modified radical neck dissection. The surgical specimen showed a 2.5 × 1.5 × 10 cm, well-circumscribed macrofollicular variant of papillary thyroid carcinoma in the right lobe and multiple central and right lateral neck lymph node metastases. Molecular testing for BRAF, NRAS, HRAS, and KRAS was all negative. We then reviewed the demographic and clinicopathologic characteristics of 71 patients with the macrofollicular variant of papillary thyroid carcinoma. The cytologic or histopathologic diagnosis of macrofollicular variant of papillary thyroid carcinoma can be difficult. Extensive lymph node metastases caused by the macrofollicular variant of papillary thyroid carcinoma may occur even in the absence of capsular or lymphovascular invasion. This review will help to better understand the nature of the macrofollicular variant of papillary thyroid carcinoma.

The prediction of malignant risk in the category “atypia of undetermined significance/follicular lesion of undetermined significance” of the Bethesda System for Reporting Thyroid Cytopathology using subcategorization and BRAF mutation results

The "atypia of undetermined significance/follicular lesion of undetermined significance" (AUS/FLUS) category in the Bethesda System for Reporting Thyroid Cytopathology is a heterogeneous category of cases that are not clearly benign or malignant. We conducted an analysis of cytologic and histologic evaluations of thyroid nodules that had been interpreted as AUS/FLUS on fine-needle aspiration (FNA) at a single institution from April 2011 to April 2012. Those cases were classified into 2 subgroups according to the predominance of nuclear atypia (AUS) or microfollicular architecture (FLUS). In addition, for a number of these cases, BRAF gene mutation analyses were performed. Of 6402 thyroid FNAs performed, 431 cases were diagnosed as AUS and 120 as FLUS. Follow-up cytologic or histologic outcome data were available for 315 AUS cases and 73 FLUS cases. Among AUS cases, 52.7% were malignant on repeat FNA or histologic diagnosis. In contrast, for FLUS, 6.8% were malignant on repeat FNA or histologic diagnosis. Among AUS/FLUS cases, 147 had adequate BRAF mutation analysis, which accompanied the histologic diagnosis. BRAF mutations were found in 87 AUS cases, 86 of which were papillary carcinoma. In contrast, there was only 1 case of BRAF mutation in FLUS. Correlating molecular results with histologic outcome revealed a 98.9% cancer probability for AUS cases with BRAF mutation. The AUS subcategory indicates a higher risk of malignancy than the FLUS subcategory. Furthermore, BRAF molecular testing is helpful in stratifying the malignant risk of AUS cases into high-risk and low-risk groups.

External quality assessment of BRAF molecular analysis in melanoma

The availability of BRAF inhibitors has given metastatic melanoma patients an effective new treatment choice and molecular testing to determine the presence or absence of a BRAF codon 600 mutation...

Diagnosis and management of differentiated thyroid cancer using molecular biology

To define molecular biology in clinical practice for diagnosis, surgical management, and prognostication of differentiated thyroid cancer. Ovid Medline 2006-2012 Manuscripts with clinical correlates. Papillary thyroid carcinomas harbor point mutations of the BRAF and RAS genes or RET/PTC rearrangements, all of which activate the mitogen-activated protein kinase pathway. These mutually exclusive mutations are found in 70% of PTC. BRAF mutation is found in 45% of papillary thyroid cancer and is highly specific. Follicular carcinomas are known to harbor RAS mutation or PAX8/PPARγ rearrangement. These mutations are also mutually exclusive and identified in 70% of follicular carcinomas. Molecular classifiers measure the expression of a large number of genes on a microarray chip providing a substantial negative predictive value pending further validation. 1) 20% to 30% of cytologically classified Follicular Neoplasms and Follicular Lesion of Undetermined Significance collectively are malignant on final pathology. Approximately 70% to 80% of thyroid lobectomies performed solely for diagnostic purposes are benign. Molecular alteration testing may reduce the number of unnecessary thyroid procedures, 2) may reduce the number of completion thyroidectomies, and 3) may lead to more individualized operative and postoperative management. Molecular testing for BRAF, RAS, RET/PTC, and PAX8/PPARγ for follicular lesion of undetermined significance and follicular neoplasm improve specificity, whereas molecular classifiers may add negative predictive value to fine needle aspiration diagnosis.

Intraoperative Pathologic Examination in the Era of Molecular Testing for Differentiated Thyroid Cancer

Diagnostic thyroidectomy is typically indicated for indeterminate thyroid cytology results. Traditionally, intraoperative pathologic examination (IOPE) helped to guide the extent of initial surgery. Preoperative molecular testing (MT) of fine needle aspiration cytology has emerged as another diagnostic adjunct, is highly specific for thyroid cancer, and can lead to appropriate initial total thyroidectomy. We hypothesized that preoperative MT obviates the need for routine IOPE during lobectomy. In a retrospective, consecutive cohort study, we compared outcomes of 670 patients undergoing thyroidectomy. Cohort A (January 2005 to December 2006) received surgery without MT, and cohort B (January 2008 to September 2010) had preoperative MT for BRAF, RAS, RET/PTC, and PAX8/PPARγ mutations, and cytology assessment by the 2007 modified Bethesda criteria. In both cohorts, IOPE was performed during lobectomy and a positive result prompted total thyroidectomy. In cohort B, total thyroidectomy was more often the initial surgery (62% vs A 45%; p < 0.001) and a positive MT result was the only factor prompting initial total thyroidectomy in 18 (9%) patients. Among 315 patients who had initial lobectomy, thyroid cancer was infrequently diagnosed by IOPE in both cohorts (A 3.6% vs B 1.7%; p = 0.5). The sensitivity of IOPE in detecting differentiated thyroid cancer ≥1 cm decreased >60% with routine use of MT and the Bethesda criteria (A 18.4% vs B 5.9%). After lobectomy, differentiated thyroid cancer ≥1 cm was equally likely to be diagnosed in both cohorts (p = 0.1), but follicular variant papillary thyroid cancer was more common in cohort B (74% vs 45%; p = 0.02). Together with the Bethesda cytologic criteria, preoperative MT allows for an increased rate of initial definitive total thyroidectomy and eliminates the need for routine intraoperative pathologic examination during diagnostic lobectomy.

Suspicious Ultrasound Characteristics Predict BRAFV600E-Positive Papillary Thyroid Carcinoma

Current American Thyroid Association (ATA) guidelines recommend routine cervical ultrasound (US) in thyroid nodule evaluation. Specific US characteristics can help diagnose papillary thyroid carcinoma (PTC). The aim of this blinded cohort study was to determine whether these specific US characteristics can also reliably detect the more aggressive variants of PTC that are often associated with the BRAF(V600E) mutation. After Institutional Review Board approval, we identified a cohort of patients from January 2007 to December 2009 with histologic PTC≥1 cm who had cervical US, initial thyroid surgery, and molecular testing for BRAF(V600E) on fine-needle aspiration biopsy or histology. Preoperative US images were evaluated by a single radiologist, who was blinded to BRAF status, for nodule size and the presence or absence of the following suspicious US features: taller-than-wide shape, ill-defined margins, hypoechogenicity, calcifications, noncystic composition, and absent halo. BRAF-positivity was associated with most known suspicious US findings, including taller-than-wide shape (47% vs. 7%, p<0.001), ill-defined margins (42% vs. 9%, p<0.001), hypoechogenicity (83% vs. 36%, p<0.001), micro/macrocalcifications (87% vs. 24%, p<0.001), and absent halo (85% vs. 27%, p<0.001) but was not associated with noncystic composition. When ≥3 suspicious US features were present, BRAF-positivity was predicted with a positive predictive value of 82%. The absence of suspicious US features together with negative BRAF testing predicted PTC without extrathyroidal extension or lymph node metastasis (negative predictive value 88%). With routine preoperative cervical US and molecular testing, a trained radiologist or surgeon can improve the preoperative characterization of PTC, potentially impacting risk stratification and initial surgical management.